[Castleman's disease in the rheumatological practice]. / Morbus Castleman in der rheumatologischen Praxis.

The term Castleman's disease encompasses a group of rare lymphoproliferative diseases that show histopathological similarities in lymph node biopsy. Diagnostic criteria and a specific ICD-10 code have been available for a few years. Case studies listed at the beginning illustrate that close cooperation between clinicians and pathologists is required to enable a reliable diagnosis. For an optimal histopathological assessment, the pathologist is also dependent on the removal of a complete lymph node. Before distinguishing a potentially fatal multicentric idiopathic Castleman's disease from the resectable unicentric form, which is important in terms of prognosis and treatment, early diagnosis presupposes that Castleman's disease is considered in the differential diagnosis. Various immune phenomena and overlaps with autoimmune diseases can increase the probability of misdiagnosis or undetected cases in the clinical routine of rheumatologists. The intention of the present overview is therefore to point out the similarities with autoimmune diseases that are relevant for differential diagnoses and to point out situations that justify a review of the previous diagnosis.

[Rebiopsy for Patients with Lung Cancer - Joint Opinion from both the Endoscopic and Thoracic Oncology Sections of the German Society of Pneumology (DGP)]. / Empfehlung der Sektionen 2 und 11 der DGP zur Rebiopsie bei Lungenkarzinomen.

Performing rebiopsies for primary lung cancer and/or their metastases is becoming more and more prominent in daily practice, as the therapeutical spectrum increases and some newer strategies are dependent on immunohistochemical and/or molecular factors. In general, nearly all recurrent lesions or metastases can be reached. However, frequently invasive procedures are necessary with the need to carefully weigh risks and benefits of rebiopsies for the patient in each case. In this review indications for recurrent and progressive disease as well as risks are discussed and alternatives to rebiopsies are shown. This work is the joint opinion from both the endoscopic and thoracic oncology sections of the German Society of Pneumology (DGP).

[Predictive PD-L1 immunohistochemistry for non-small cell lung cancer : Current state of the art and experiences of the first German harmonization study]. / Prädiktive PD-L1-Immunhistochemie beim nichtkleinzelligen Bronchialkarzinom : Aktueller Stand und Erfahrungen der ersten deutschen Harmonisierungsstudie.

BACKGROUND: Antibodies against PD-1 and PD-L1 can cause strong and durable anti-tumor immune responses in non-small cell lung cancer (NSCLC). Immunohistochemistry for PD-L1 (PD-L1 IHC) was tested as a predictive biomarker. Several IHC assays and interpretation criteria were developed in parallel. AIM: The clinical significance of PD-L1 IHC in NSCLC and the optimum method for staining and interpretation of the results are the subject of ongoing studies. The diagnostic application of immunotherapy in NSCLC necessitates harmonization of PD-L1 IHC to obtain evidence for guidelines; therefore, a consensus opinion on a well-founded diagnostic mode of testing should be defined based on published studies and the results of the first German PD-L1 IHC harmonization study. METHODS: 1. Summary of the current data situation. 2. Evaluation of the first German PD-L1 IHC harmonization study (centralized, staining with PD-L1 IHC analogous to studies, 15 cases of NSCLC, 4 IHC study assays [28­8, 22C3, SP142 and SP263] and scoring by 9 pathologists). RESULTS: The use of PD-L1 IHC in NSCLC is suitable for identification of patients with an increased probability of a clinical benefit from immunotherapy. The various proportional cut-offs used to interpret the staining results can be summarized in a total score, which can be reproducibly assessed. The staining patterns of the four assays investigated were, however, not congruent in all situations. DISCUSSION: In principle, the use of PD-L1 IHC for assessment of the expression in tumor cells is a reliably determinable biomarker. Evaluation algorithms should be based on published clinical trials. For NSCLC approvals with obligatory PD-L1 IHC are to be expected but it remains to be seen to what extent PD-L1 IHC will be implemented in the clinical routine.

[Sport as a means to prevention, rehabilitation, and health promotion. An expert opinion]. / Sport als Mittel in Prävention, Rehabilitation und Gesundheitsförderung. Eine Expertise.

Physical activities are particularly health effective when they are structured and systematically organized, for example, with respect to a dose-response relationship or to psychosocial aspects. Which insights can be gained from scientific knowledge on the structure of physical activities? Which health-related goals can be sought with different physical activities in the field of sport? Which target groups can be reached with these kinds of sport? These questions are addressed in this article on the basis of current scientific knowledge-to promote the development of health-enhancing physical activities especially in sport federations, but also to create a communication platform between the sport and the health sector. The work of researchers in this field is focused mainly on health-enhancing physical activity, but they are also engaged in health-related scientific organizations and in sport federations.

Epstein-Barr virus-associated diffuse large B-cell lymphoma of the hypopharynx.

PROBLEMS/OBJECTIVES: Epstein-Barr virus (EBV) is commonly associated with nasopharyngeal carcinoma and Burkitt's lymphoma, but association with hypopharyngeal and laryngeal tumours is rare. To the best of our knowledge, this is the first case report of an EBV-associated diffuse large B-cell lymphoma (DLBCL) of the hypopharynx. METHODOLOGY: A 63-year-old male patient suffering from chronic lymphocytic leukemia presented with swallowing disorders and a sore throat. Panendoscopy with laser surgical resection of tissue specimens was performed. RESULTS: Immunohistochemical and molecular genetic diagnostics, including EBV-encoded small RNA in situ hybridization, confirmed the diagnosis of an EBV-associated DLBCL of the hypopharynx. Ten weeks after the diagnosis, the patient died of disease related to multiple complications. CONCLUSIONS: We hypothesize that the EBV infection was triggered by long-term immunosuppressive therapy that led secondarily to the development of a DLBCL. Otorhinolaryngologists should keep in mind that lymphomas might develop in the entire pharynx.

Immune related endonucleases and GTPases are not associated with tumor response in patients with advanced non-small cell lung cancer treated with checkpoint inhibitors.

Immune related endonucleases have recently been described as potential therapeutic targets and predictors of response to treatment with immune checkpoint inhibitors (ICI). The aim is to evaluate the association between the expression of 5 biomarkers involved in the immune response (CD73, CD39, VISTA, Arl4d and Cytohesin-3) in parallel with the more common ICI-predictive markers, PD-L1 expression and Tumor Mutation Burden (TMB) with response to ICI therapy in an advanced non-small cell lung cancer (NSCLC) cohort. METHODS: Patients with advanced NSCLC treated with ICI single agent were divided into responders and non-responders according to RECIST v1.1 and duration of response (DOR) criteria. Immunohistochemistry was performed on pretreatment tumor tissue samples for PD-L1, CD73, CD39, VISTA, Arl4d, and Cytohesin-3 expression. TMB was estimated with NEOplus v2 RUO (NEO New Oncology GmbH) hybrid capture next generation sequencing assay. Resistance mutations in STK11/KEAP1 and positive predictive mutations in ARID1A/POLE were also evaluated. RESULTS: Included were 56 patients who were treated with ICI single agent. The median progression-free and overall survival for the whole cohort was 3.0 (95% CI, 2.4-3.6) and 15 (95% CI, 9.7-20.2) months, respectively. The distribution of CD73 in tumor cells and CD39, VISTA, Arl4d and Cytohesin-3 expression in immune cells were not different between responders and non-responders. Also, PD-L1 and TMB were not predictive for response. The frequency of STK11, KEAP1 and ARID1A mutations was low and only observed in the non-responder group. CONCLUSION: Separate and combined expression of 5 biomarkers involved in the immune response (CD73, CD39, VISTA, Arl4d, and Cytohesin-3) was not associated with response in our cohort of advanced NSCLC patients receiving single agent ICI. To confirm our findings the analysis of independent larger cohorts is warranted.

Combined histochemical approach in assessing tryptase expression in the mast cell population.

To increase the efficiency of interpretation of mast cell's contribution to the state of a specific tissue microenvironment, it is necessary to detail the molecular composition of their secretome and analyze the pathways of degranulation. Developed method of combining immunomorphological and histochemical staining protocols contributes to the most objective detection of the integral level of tryptase expression in the intraorgan population of the skin mast cells. Novel technique for tryptase detection expands the possibilities of morphological analysis, provides researchers with additional data on the structure of the mast cell population and helps visualize the processing and cytological features and structural targets of tryptase during the development of adaptive and pathological reactions. Objective determination of the tryptase profile for organ-specific mast cell populations is in great demand in clinical practice for the interpretation of pathological processes, including inflammation and oncogenesis.

Powerful strategy for polymerase chain reaction-based clonality assessment in T-cell malignancies Report of the BIOMED-2 Concerted Action BHM4 CT98-3936.

Polymerase chain reaction (PCR) assessment of clonal T-cell receptor (TCR) and immunoglobulin (Ig) gene rearrangements is an important diagnostic tool in mature T-cell neoplasms. However, lack of standardized primers and PCR protocols has hampered comparability of data in previous clonality studies. To obtain reference values for Ig/TCR rearrangement patterns, 19 European laboratories investigated 188 T-cell malignancies belonging to five World Health Organization-defined entities. The TCR/Ig spectrum of each sample was analyzed in duplicate in two different laboratories using the standardized BIOMED-2 PCR multiplex tubes accompanied by international pathology panel review. TCR clonality was detected in 99% (143/145) of all definite cases of T-cell prolymphocytic leukemia, T-cell large granular lymphocytic leukemia, peripheral T-cell lymphoma (unspecified) and angioimmunoblastic T-cell lymphoma (AILT), whereas nine of 43 anaplastic large cell lymphomas did not show clonal TCR rearrangements. Combined use of TCRB and TCRG genes revealed two or more clonal signals in 95% of all TCR clonal cases. Ig clonality was mostly restricted to AILT. Our study indicates that the BIOMED-2 multiplex PCR tubes provide a powerful strategy for clonality assessment in T-cell malignancies assisting the firm diagnosis of T-cell neoplasms. The detected TCR gene rearrangements can also be used as PCR targets for monitoring of minimal residual disease.

HMG-CoA reductase inhibitors (statins) increase endothelial progenitor cells via the PI 3-kinase/Akt pathway.

HMG-CoA reductase inhibitors (statins) have been developed as lipid-lowering drugs and are well established to reduce morbidity and mortality from coronary artery disease. Here we demonstrate that statins potently augment endothelial progenitor cell differentiation in mononuclear cells and CD34-positive hematopoietic stem cells isolated from peripheral blood. Moreover, treatment of mice with statins increased c-kit(+)/Sca-1(+)--positive hematopoietic stem cells in the bone marrow and further elevated the number of differentiated endothelial progenitor cells (EPCs). Statins induce EPC differentiation via the PI 3-kinase/Akt (PI3K/Akt) pathway as demonstrated by the inhibitory effect of pharmacological PI3K blockers or overexpression of a dominant negative Akt construct. Similarly, the potent angiogenic growth factor VEGF requires Akt to augment EPC numbers, suggesting an essential role for Akt in regulating hematopoietic progenitor cell differentiation. Given that statins are at least as potent as VEGF in increasing EPC differentiation, augmentation of circulating EPC might importantly contribute to the well-established beneficial effects of statins in patients with coronary artery disease.

[Coagulation and formation of malignant effusions]. / Die Rolle des Gerinnungssystems bei der Entstehung maligner Ergüsse.

Malignant effusions are a frequent problem for cancer patients. Due to the high resistance of tumor cells within these effusions, no effective treatment has been defined yet. Most patients exhibit additional phenomena related to hyper-coagulability such as elevated levels for d-dimers and prothrombin fragments f1.2; half of them suffer from manifest thrombosis or complications. We followed the hypothesis that the activated coagulation system contributes to the resistance of tumor cells and analyzed the effusions from cancer patients. The majority of isolated tumor cells aberrantly expressed PAR-1 thrombin receptors. In vitro pre-incubation of PAR-1 expressing human leukemia cells with thrombin resulted in a dose-dependent resistance to idarubicin. Within the effusions, we did not only find high concentrations of VEGF and tissue factor, but also all coagulation factors of the tissue factor pathway. Very high levels of prothrombin fragments f1.2 indicate constant thrombin generation. Upon the basis of these findings, we developed a multistep model elucidating the pathophysiological generation of malignant effusions, which might serve as a basis for further examinations.

Primary mediastinal large B-cell lymphoma with sclerosis in pediatric and adolescent patients: treatment and results from three therapeutic studies of the Berlin-Frankfurt-Münster Group.

PURPOSE: Primary mediastinal large B-cell lymphoma with sclerosis (PMLBL) is a rare entity of non-Hodgkin's lymphoma (NHL) arising from thymic mature B cells. Optimal treatment strategies remain to be established, especially in pediatric patients. PATIENTS AND METHODS: This study analyzes clinical characteristics and treatment outcome of 30 pediatric patients with PMLBL, diagnosed in multicenter therapy NHL-Berlin-Frankfurt-Münster Group (BFM) trials. Treatment was stratified by stage and serum lactate dehydrogenase (LDH) and consisted of four to six 5-day courses of chemotherapy using steroids, oxazaphosphorine alkylating agents, methotrexate, cytarabine, etoposide, and doxorubicin. Radiation was not part of the protocol. RESULTS: From April 1986 to August 1999, 1,650 patients with newly diagnosed NHL were enrolled in the NHL-BFM trials; 30 patients (1.8%) had PMLBL. Median age was 14.3 years (range, 1.4 to 16.7 years); 15 patients were male and 15 patients were female. With a median observation time of 5 years (range, 1 to 12 years), probability of event-free survival (pEFS) at 5 years was 0.70 (SE, 0.08). Two patients erroneously diagnosed as T-cell NHL received non-B-cell therapy and died from progress of disease. Events in 28 patients receiving B-cell therapy included early progress during therapy (n = 1) and relapse (n = 6). Residual mediastinal masses were present in 23 patients after two courses of therapy and in 15 patients after the end of therapy. LDH > or = 500 U/L was associated with increased risk of failure in multivariate analysis. CONCLUSION: PMLBL mainly is found in adolescents. Dose-intense chemotherapy including high-dose methotrexate yields a pEFS at 5 years of 0.70 (SE, 0.08). LDH is of prognostic value in pediatric patients with PMLBL.

Successful treatment strategy for Ki-1 anaplastic large-cell lymphoma of childhood: a prospective analysis of 62 patients enrolled in three consecutive Berlin-Frankfurt-Munster group studies.

PURPOSE: To prove prospectively the efficacy of a short-pulse chemotherapy for treatment of Ki-1 anaplastic large-cell lymphoma (ALCL) of childhood. PATIENTS AND METHODS: From October 1983 to December 1992, 62 patients (median age, 9.7 years) with newly diagnosed Ki-1 ALCL were enrolled onto Non-Hodgkin's Lymphoma-Berlin-Frankfurt-Munster (NHL-BFM) studies 83, 86, and 90. The most frequent immunophenotype was T cell. Ki-1 ALCL differed from other subsets of NHL of childhood by the more frequent involvement of bone, soft tissue, and skin, and by the lack of bone marrow (BM) disease. A 5-day prephase course (prednisone/cyclophosphamide) was followed by two different 5-day courses of chemotherapy: course A consisted of dexamethasone, methotrexate (MTX) 0.5 g/m2 (24-hour infusion), intrathecal chemotherapy, ifosfamide, cytarabine (Ara-C), and etoposide (VP-16); course B consisted of cyclophosphamide and doxorubicin instead of ifosfamide, and Ara-C/VP-16, respectively. Treatment was stratified into three branches. Branch 1 (stage I and stage II resected) received three courses; branch 2 (stage II not resected, stage III), six courses; and branch 3 (stage IV), six intensified courses containing MTX 5 g/m2, and Ara-C 2 g/m2. Local radiotherapy was not performed. RESULTS: Four patients failed to enter remission, and one died of infection. Seven patients relapsed within 9 months after diagnosis; two patients had isolated local relapses, but BM and CNS were never involved. Fifty patients have been in first continuous complete remission (CR) for 0.6 to 9.7 years (median, 2.5), and 56 are alive. The probabilities for survival and event-free survival (EFS) at 9 years are 83% +/- 7% (SE) and 81% +/- 5%. Skin involvement was the only negative prognostic parameter. CONCLUSION: Short-pulse chemotherapy over 2 to 5 months without local therapy modalities is effective in the treatment of Ki-1 ALCL.

Mantle-cell lymphomas have more widespread disease and a slower response to chemotherapy compared with follicle-center lymphomas: results of a prospective comparative analysis of the German Low-Grade Lymphoma Study Group.

PURPOSE: To compare mantle-cell lymphomas (MCLs) and follicle-center lymphomas (FCLs) for their features of clinical presentation, response to chemotherapy, and prognosis on the basis of a prospective randomized clinical trial. PATIENTS AND METHODS: Patients with MCL and FCL who entered onto the prospective randomized comparison of cyclophosphamide, vincristine, and prednisone (COP) versus prednimustine and mitoxantrone (PmM) followed by a second randomization for interferon (IFN) maintenance versus observation only. RESULTS: One hundred sixty-five of 234 patients had FCL and 45 of 234 patients had MCL. With FCL, both sexes were equally affected (men, 47%); patients with MCL were predominantly men (78%; P < .0004) and had a higher median age (64 v 53 years; P < .0001). Patients with MCL also had more widespread disease, reflected by the proportion of patients with two or greater extranodal manifestations (43% v 21%; P < .005) and nine or greater involved nodal areas (64% v 45%; nonsignificant [NS]). Response to chemotherapy was significantly lower in patients with MCL (complete remission [CR] + partial remission [PR], 69% v 88%; P < .05) and occurred at a slower pace. Patients with MCL also had a shorter event-free interval (median, 8 v 24 months; P < .0001) and overall survival (median, 28 v 77 months; P < .0001). In both subtypes, however, patients with less than two residual lymphoma manifestations in remission experienced a relatively good prognosis with an estimated 5-year survival of greater than 60% for MCL and greater than 75% for FCL. CONCLUSION: MCL and FCL differ substantially in their features of presentation, response to chemotherapy, and long-term prognosis. The extent of residual disease after completion of chemotherapy discriminates patients with different prognosis and may be used for the stratification of postremission strategies.

Primary gastrointestinal non-Hodgkin's lymphoma: I. Anatomic and histologic distribution, clinical features, and survival data of 371 patients registered in the German Multicenter Study GIT NHL 01/92.

PURPOSE: The study was initiated to obtain epidemiologic data and information on anatomic and histologic distribution, clinical features, and treatment results in patients with primary gastrointestinal non-Hodgkin's lymphomas (PGI NHL). PATIENTS AND METHODS: Between October 1992 and November 1996, 371 PGI NHL patients were eligible to evaluate clinical features. Radiotherapy and chemotherapy were stratified according to histologic grading, stage, and whether surgery had been carried out or not. RESULTS: A total of 74.8% patients had gastric NHL (PGL). Within the intestine, the small bowel and the ileocecal region were involved in 8.6% and 7.0% of the cases, respectively. Multiple GI involvement (MGI) was 6.5%. Approximately 90% of the GI NHL were in stages IE/IIE. Aggressive NHL accounted for the majority, with a distinguishable pattern in several sites. Forty percent of PGL were of low-grade mucosa-associated lymphatic tissue type. One third of large-cell lymphomas had low-grade components. Most intestinal NHL were germinal-center lymphomas. The site of origin was prognostic. In gastric and ileocecal lymphoma, event-free (EFS) and overall survival (OS) were significantly higher as compared with the small intestine or MGI (median time of observation, 51 months). In PGL, localized disease was prognostic for EFS and OS. Histologic grade influenced only EFS significantly. Numbers in intestinal lymphomas were too small for subanalyses. CONCLUSION: PGI NHL are heterogeneous diseases. The number of localized PGL allowed for detailed analyses. Larger studies are needed for stages III and IV and for intestinal NHL. A uniform reporting system for PGI NHL, in terms of definitions and histologic and staging classifications, is needed to facilitate comparison of treatment results.

Primary gastrointestinal non-Hodgkin's lymphoma: II. Combined surgical and conservative or conservative management only in localized gastric lymphoma--results of the prospective German Multicenter Study GIT NHL 01/92.

PURPOSE: The aim of the study was to obtain data on anatomic and histologic distribution, clinical features, and treatment results of patients with primary gastrointestinal non-Hodgkin's lymphomas, particularly combined surgical and conservative treatment (CSCT) versus conservative treatment (CT) alone for primary gastric lymphoma (PGL) in localized stages. PATIENTS AND METHODS: Whether the treatment included surgery was left to the discretion of each participating center. Radiotherapy (Rx) and chemotherapy were stratified according to histologic grading, stage, and the inclusion or omission of surgery as follows: patients with low-grade PGL were treated with extended-field (EF) Rx (30 Gy). In case of residual tumor after surgery or in case of CT only (in stage IIE after six cycles of cyclophosphamide, vincristine, and prednisone), an additional boost of 10 Gy was given. All patients with high-grade PGL were treated with four (stage IE) or six (stage IIE) cycles of cyclophosphamide, doxorubicin, vincristine, and prednisone followed by EF Rx (stage IE) or involved-field (IF) Rx (stage IIE). Rx dosage corresponded to low-grade NHL. RESULTS: Between October 1992 and November 1996, 106 patients had CT only. The survival rate (SR) after 5 years was 84.4% and was influenced neither by patients' characteristics nor by stage or histologic grade. Seventy-nine patients had CSCT. Their SR was 82.0%. Complete resection of the tumor (R0) was prognostic for the overall survival (P =.0165) as compared with incomplete resection. CONCLUSION: Although the study was not randomized, a stomach-conserving approach may be favored.

Sequential application of chemotherapy and monoclonal CD 20 antibody: successful treatment of advanced composite-lymphoma.

We describe successful treatment of a 38-year-old patient with composite lymphoma stage IV(A), who presented with multifocal enlarged lymph nodes. The lymph node histology showed classic morphologic features of Hodgkin's disease, mixed cellularity subtype and follicular B-cell lymphoma. Immunophenotypic analysis showed immunoreactivity for CD20, CD10 and Ki-67 in the malignant small cell population. The areas of Hodgkin's disease demonstrated positive immunoreactivity for CD30 and CD20 in the Hodgkin's cells. Both cell populations were bcl2-oncoprotein positive. Eight courses of dose-escalated BEACOPP were administered. Restaging after chemotherapy showed radiological partial remission, but biopsy confirmed persisting follicular B-cell lymphoma without bone marrow infiltration and no evidence of Hodgkin's disease. He was treated with monoclonal CD 20-antibody (Rituximab) 10 mg/kg weekly for eight consecutive weeks due to marked positivity of CD 20-antigen in follicular lymphoma cells. This treatment was well tolerated and final staging showed complete remission of the composite lymphoma. This patient continues to be in remission 28 months after the end of the treatment. In conclusion, in the very rare case of composite lymphoma a combination of chemotherapy and subsequent immunotherapy might be considered as a promising therapeutic option.

Identification and structural analysis of rearranged immunoglobulin heavy chain genes in lymphomas and leukemias.

The third complementarity determining region (CDR3) of the hypervariable domain of immunoglobulin heavy chain (IgH) genes represents a highly variable and clone-specific IgH-CDR3 sequences in 10 non-Hodgkin's lymphomas (NHL), five chronic lymphocytic leukemias (CLL) and five acute lymphoblastic leukemias (ALL) of B cell lineage. The IgH-CDR3 sequences were amplified using DNA extracted from clinical specimens (bone marrow, peripheral blood and fresh-frozen or paraffin-embedded lymph nodes) by a semi-nested PCR with consensus primers directed to conserved regions within the variable (VH) and the joining (JH) gene segments. In 17/20 samples (85%), a distinct IgH-CDR3 PCR product was obtained. Individual PCR products were sequenced after cloning. The nucleotide sequences of 134 randomly chosen recombinant vectors were determined demonstrating in 17/20 cases (85%) monoclonal VH-N-DH-N-JH junctions. Analysis of PCR products by temperature-gradient gel electrophoresis (TGGE) confirmed the specificity of the IgH-CDR3 PCR/sequencing results. Moreover, the combination of PCR/TGGE technology allowed the rapid and specific characterization of clonal IgH-CDR3 junctions in B cell proliferations by direct sequencing even in the presence of admixed polyclonal B cells.

Topoisomerase IIalpha expression in mantle cell lymphoma: a marker of cell proliferation and a prognostic factor for clinical outcome.

Mantle cell lymphoma (MCL) is a malignant lymphoma associated with a relatively aggressive clinical course and a median overall survival time of 3-4 years. Treatment usually consists of combination chemotherapy, often including topoisomerase (topo) inhibitors such as doxorubicin, etoposide and mitoxantrone. Topo IIalpha is an enzyme that is needed whenever uncoiling of DNA is necessary during the cell cycle. The enzyme is a marker of cell proliferation. We analyzed the expression of topo IIalpha in relation to Ki-67 and the clinical outcome in patients with MCL. Biopsy specimens from 95 untreated patients enrolled in two multicenter trials (1975-1985) were investigated immunohistochemically with monoclonal antibodies against topo IIalpha (Ki-S4) and Ki-67 (Ki-S5). Patients with low (0-10%) topo IIalpha expression had a median overall survival time of 49.0 months, compared to 17.0 months for patients with high (more than 10%) topo IIalpha expression. The Kaplan-Meier analysis showed a significant difference in the overall survival time related to the percentage of topo IIalpha (P<0.001) and Ki-67 (P<0.001) positive tumor cells. Multivariate Cox regression analysis revealed the expression of topo IIalpha as the most important prognostic factor (P<0.001) in MCL superior to the international prognostic index (IPI), the Ki-67 index and other clinical characteristics.

Structure of Bcl-1 and IgH-CDR3 rearrangements as clonal markers in mantle cell lymphomas.

Mantle cell lymphoma represent a clinicopathologically distinct entity of malignant non-Hodgkin's lymphoma (NHL) and are characterized by a specific chromosomal translocation t(11;14)(q13;q32) involving the cyclin D1 gene also designated as bcl-1/PRAD1 gene on chromosome 11 and the heavy chain immunoglobulin joining region on chromosome 14. We have established a PCR method to amplify t(11;14) junctional sequences in DNA from fresh frozen and paraffin-embedded tissue by bcl-1-specific primers in combination with a consensus immunoglobulin JH primer. A total of 65 cases histologically classified as mantle cell lymphoma (MCL) were analyzed for the presence of a t(11;14) translocation and monoclonal IgH-CDR3 rearrangements. From 26 patients with classical MCL and three cases with the anaplastic variant of MCL fresh frozen biopsy material was available for DNA extraction. We detected a bcl-1/JH rearrangement in 12 out of 29 samples (41%). In 36 cases paraffin-embedded lymph node tissue was the only source of DNA. In this material we found a bcl-1/JH rearrangement in six out of 31 samples with intact DNA (20%). To confirm the specificity of the PCR and to determine the bcl-1/JH junctional region sequences as clone-specific marker in individual patients we characterized the junctional DNA sequences by direct PCR sequencing in 16 cases. Interestingly we found that six bcl-1/JH junctions harbored DH segments in their N regions indicating that bcl-1/JH rearrangements can occur in a later stage of B cell ontogeny during which the complete VH to DH-JH joining or VH-replacement takes place. To investigate the suitability of IgH-CDR3 as sensitive molecular marker for those MCL patients in which a t(11;14) translocation can not easily be amplified, we additionally analysed 60 cases for the presence of monoclonally rearranged IgH genes by IgH-CDR3-PCR. A monoclonal IgH-CDR3 PCR product could be identified in 24 out of 29 fresh frozen samples (79%) whereas only 11 out of 31 samples (36%) with paraffin-derived DNA were positive. We demonstrate that automated fluorescence detection of monoclonal IgH-CDR3 PCR products allows the rapid and sensitive monitoring of minimal residual disease also in cases that lack a PCR amplifiable t(11;14) translocation. In combination with allele-specific primers the procedure may improve current experimental approaches for detection of occult MCL cells at initial staging and residual disease during and after therapy.

Altered expression of the retinoblastoma gene product in human high grade non-Hodgkin's lymphomas.

The retinoblastoma gene (RB) is a growth suppressor gene on the human chromosome 13q14. It encodes a 105 kDa phosphoprotein (p105), with DNA-binding capacity. P105 is thought to be involved in cell cycle control. Inactivation of RB is responsible for the development of retinoblastomas and occurs frequently in osteosarcomas and small cell lung cancer. In this study we looked at the RB-structure and expression in cell lines and primary lymphoma samples from patients with high grade non-Hodgkin's lymphoma (NHL). Forty five primary high grade NHL, the B-lymphoblastoid cell line IM-9 and the NHL cell line WSU-NHL were studied for RB structure by Southern blotting and for RB-expression by Northern blotting, Western blotting and immunocytochemistry. In all experiments freshly cryopreserved material was used. Southern and Northern experiments were performed with the 0.9 kb and 3.8 kb RB-cDNA probe. For the detection of p105 two different anti-p105-monoclonal antibodies were used in immunocytochemistry and Western blotting experiments. No RB mRNA and no p105 could be found in IM-9 cells. Twenty six high grade NHL samples (58%) showed no p105 expression. In the subgroup of centroblastic lymphomas 16 out of 21 and in Burkitt's lymphomas five out of eight showed no p105-expression. P105 expression is absent in 58% of high grade NHL, particularly in centroblastic and Burkitt's lymphomas, suggesting that inactivation of RB may play a crucial role in the pathogenesis of high grade NHL.